Cytokine responses in nonlesional psoriatic skin as clinical predictor to anti-TNF agents.

BACKGROUND: A major issue with the current management of psoriasis is our inability to predict treatment response. OBJECTIVE: Our aim was to evaluate the ability to use baseline molecular expression profiling to assess treatment outcome for patients with psoriasis. METHODS: We conducted a longitudinal study of 46 patients with chronic plaque psoriasis treated with anti-TNF agent etanercept, and molecular profiles were assessed in more than 200 RNA-seq samples. RESULTS: We demonstrated correlation between clinical response and molecular changes during the course of the treatment, particularly for genes responding to IL-17A/TNF in keratinocytes. Intriguingly, baseline gene expressions in nonlesional, but not lesional, skin were the best marker of treatment response at week 12. We identified USP18, a known regulator of IFN responses, as positively correlated with Psoriasis Area and Severity Index (PASI) improvement (P = 9.8 × 10-4) and demonstrate its role in regulating IFN/TNF responses in keratinocytes. Consistently, cytokine gene signatures enriched in baseline nonlesional skin expression profiles had strong correlations with PASI improvement. Using this information, we developed a statistical model for predicting PASI75 (ie, 75% of PASI improvement) at week 12, achieving area under the receiver-operating characteristic curve value of 0.75 and up to 80% accurate PASI75 prediction among the top predicted responders. CONCLUSIONS: Our results illustrate feasibility of assessing drug response in psoriasis using nonlesional skin and implicate involvement of IFN regulators in anti-TNF responses.

Neuroprotective effects of long noncoding RNAs involved in ischemic postconditioning after ischemic stroke.

During acute reperfusion, the expression profiles of long noncoding RNAs in adult rats with focal cerebral ischemia undergo broad changes. However, whether long noncoding RNAs are involved in neuroprotective effects following focal ischemic stroke in rats remains unclear. In this study, RNA isolation and library preparation was performed for long noncoding RNA sequencing, followed by determining the coding potential of identified long noncoding RNAs and target gene prediction. Differential expression analysis, long noncoding RNA functional enrichment analysis, and co-expression network analysis were performed comparing ischemic rats with and without ischemic postconditioning rats. Rats were subjected to ischemic postconditioning via the brief and repeated occlusion of the middle cerebral artery or femoral artery. Quantitative real-time reverse transcription-polymerase chain reaction was used to detect the expression levels of differentially expressed long noncoding RNAs after ischemic postconditioning in a rat model of ischemic stroke. The results showed that ischemic postconditioning greatly affected the expression profile of long noncoding RNAs and mRNAs in the brains of rats that underwent ischemic stroke. The predicted target genes of some of the identified long noncoding RNAs (cis targets) were related to the cellular response to ischemia and stress, cytokine signal transduction, inflammation, and apoptosis signal transduction pathways. In addition, 15 significantly differentially expressed long noncoding RNAs were identified in the brains of rats subjected to ischemic postconditioning. Nine candidate long noncoding RNAs that may be related to ischemic postconditioning were identified by a long noncoding RNA expression profile and long noncoding RNA-mRNA co-expression network analysis. Expression levels were verified by quantitative real-time reverse transcription-polymerase chain reaction. These results suggested that the identified long noncoding RNAs may be involved in the neuroprotective effects associated with ischemic postconditioning following ischemic stroke. The experimental animal procedures were approved by the Animal Experiment Ethics Committee of Kunming Medical University (approval No. KMMU2018018) in January 2018.

Negative regulation by proBDNF signaling of peripheral neurogenesis in the sensory ganglia of adult rats.

Neurogenesis in the adult brain is well recognized and plays a critical role in the maintenance of brain function and homeostasis. However, whether neurogenesis also occurs in the adult peripheral nervous system remains unknown. Here, using sensory ganglia (dorsal root ganglia, DRGs) as a model, we show that neurogenesis also occurs in the peripheral nervous system, but in a manner different from that in the central nervous system. Satellite glial cells (SGCs) express the neuronal precursor markers Nestin, POU domain, class 4, transcription factor 1, and p75 pan-neurotrophin receptor. Following sciatic nerve injury, the suppression of endogenous proBDNF by proBDNF antibodies resulted in the transformation of proliferating SGCs into doublecortin-positive cells in the DRGs. Using purified SGCs migrating out from the DRGs, the inhibition of endogenous proBDNF promoted the conversion of SGCs into neuronal phenotypes in vitro. Our findings suggest that SGCs are neuronal precursors, and that proBDNF maintains the SGC phenotype. Furthermore, the suppression of proBDNF signaling is necessary for neuronal phenotype acquisition by SGCs. Thus, we propose that peripheral neurogenesis may occur via the direct conversion of SGCs into neurons, and that this process is negatively regulated by proBDNF.

Salidroside inhibits NLRP3 inflammasome activation and apoptosis in microglia induced by cerebral ischemia/reperfusion injury by inhibiting the TLR4/NF-κB signaling pathway.

BACKGROUND: The NOD-like receptor family pyrin domain-containing 3 (NLRP3) inflammasome is an important mediator of neuroinflammatory responses that regulates inflammatory injury following cerebral ischemia and may be a potential target. Salidroside (Sal) has good anti-inflammatory effects; however, it remains unclear whether Sal can regulate NLRP3 inflammasome activation through the Toll-like receptor 4 (TLR4)/nuclear factor kappa B (NF-κB) signaling pathway after cerebral ischemia to alleviate inflammatory injury. METHODS: We established an oxygen-glucose deprivation and reoxygenation (OGD/R) model of BV2 cells and a middle cerebral artery occlusion/reperfusion (MCAO/R) rat model. Cell Counting Kit-8 (CCK-8), flow cytometry and terminal deoxynucleotidyl transferase dUTP nick-end labeling (TUNEL) assay were used to detect the viability and apoptosis of BV2 cells. Enzyme-linked immunosorbent assay (ELISA) was used to detect the level of inflammatory factors. 2,3,5-triphenyltetrazolium chloride (TTC) staining and modified Neurological Severity Score (mNSS) were used to detect cerebral infarction volume and neurological deficit in rats. Western blot, immunohistochemistry and immunofluorescence staining were used to detect the protein expression levels. RESULTS: Our results showed that Sal increased viability, inhibited lactate dehydrogenase (LDH) release, and reduced apoptosis in OGD/R-induced BV2 cells. Sal reduced the levels of tumor necrosis factor-α (TNF-α), interleukin (IL)-6, and IL-8. Following induction by OGD/R, BV2 cells exhibited NLRP3 inflammasome activation and increased protein levels of NLRP3, apoptosis-associated speck-like protein containing a CARD (ASC), caspase-1, IL-1ß, and IL-18. Protein levels of key TLR4 signaling pathway elements, such as TLR4, myeloid differentiation primary response 88 (MyD88), and phosphorylated nuclear factor kappa B p65 (p-NF-κB p65)/NF-κB p65 were upregulated. Interestingly, it was revealed that Sal could reverse these changes. In addition, TAK242, a specific inhibitor of TLR4, had the same effect as Sal treatment on BV2 cells following induction by OGD/R. In the MCAO/R rat model, Sal was also observed to inhibit NLRP3 inflammasome activation in microglia, reduce cerebral infarction volume, and inhibit apoptosis. CONCLUSIONS: In summary, we found that Sal inhibited NLRP3 inflammasome activation and apoptosis in microglia induced by cerebral ischemia/reperfusion injury by inhibiting the TLR4/NF-κB signaling pathway, thus playing a protective role. Therefore, Sal may be a promising drug for the clinical treatment of ischemic stroke.

High bonding strength between zirconia and composite resin based on combined surface treatment for dental restorations.

Zirconia is the preferred material for dental restorations; however, dental restorations are usually affected by zirconia fractures due to chipping and delamination of the veneer ceramic. One effective solution for repairing chemically inert zirconia frameworks is to strongly chemically bond them with the composite resin via surface modification. Thus, the bonding strength between the zirconia and composite resin determines the performance of dental restoration. Herein, we investigate the shear bond strength between zirconia ceramic and two ceramic repair systems before and after thermal cycling based on different surface pretreatments, including air-abrasion and a novel silane coupling agent. When treated with combined sandblasting, novel silane and 10-methacryloyloxydecyl hydrogen phosphate act as a bonding agent for the zirconia surface, and the maximum shear bond strength achieves 27.5 MPa, as measured by a universal testing machine through the average of 16 separate measurements. The results show that the combined treatment resists the interface damage caused by expansion and contraction during thermal cycling. The long-term bond durability is due to the micro-mechanical bond force formed by resin and ceramic, and the chemical bonds of Zr-O-Si at the interface. Results indicate that selective pretreating the surface results in high bond strength between the zirconia and the composite resin, which is meaningful to optimize dental restoration.

Preparation and In Vitro/Vivo Evaluation of Nano-Liposomal Form of Febrifugine Hydrochloride.

Febrifugine hydrochloride (FFH) has strong pharmacological antimalarial effect. However, compared with oral administration, the efficacy of intravenous administration is significantly reduced. In this study, we prepared conventional liposomes and PEGylated liposomes to improve the efficacy of its intravenous injection. Both liposome formulations were prepared using a modified ethanol injection method. Their mean particle sizes were 126.23 and 114.93 nm, mean zeta potentials were -6.25 and -26.33 mV, and entrapment efficiencies (EE) were 89.43 and 96.42%, respectively. The in vitro release profile indicated that the release of FFH from PEGylated liposomes and conventional liposomes was slower than free FFH, with sustained-release effect of PEGylated liposomes being more significant. PEGylated liposomes demonstrated excellent antimalarial activities in vitro superior to free FFH and conventional FFH-loaded liposomes. In addition, the PEGylated liposomes resulted in enhanced antimalarial effect in P. berghei infected mice in vivo with delayed recrudescence and prolonged survival time, compared with free FFH and conventional FFH-loaded liposomes administration. Based on these exciting experimental results, PEGylated liposomes could be a potential drug delivery system for FFH, with enhanced pharmacodynamics of intravenous injection.

Enhanced bonding strength between lithium disilicate ceramics and resin cement by multiple surface treatments after thermal cycling.

All-ceramic restoration has become a popular technology for dental restoration; however, the relative bond strength between the ceramic and resin limits its further application. Long-term high bond strength, especially after thermal cycling, is of great importance for effective restoration. The effect of physical and/or chemical surface treatments on bonding durability is seldom reported. To overcome this problem, we investigate the bond strength between lithium disilicate ceramics (LDC) and two kinds of resin cements before and after thermal cycling for a variety of surface treatments including hydrofluoric acid, two kinds of silane and a combined effect. The shear bond strength in every group is characterized by universal mechanical testing machine averaged by sixteen-time measurements. The results show that when treated with HF and a mixed silane, the LDC surface shows maximum bonding strengths of 27.1 MPa and 23.3 MPa with two different resin cements after 5000 thermal cycling, respectively, indicating an excellent ability to resist the damage induced by cyclic expansion and contraction. This long-term high bond strength is attributed to the combined effect of micromechanical interlocking (physical bonding) and the formation of Si-O-Si and -C-C- at the interface (chemical bonding). This result offers great potential for enhancing bond strength for all-ceramic restoration by optimizing the surface treatment.

3D Printing Technology in Planning Thumb Reconstructions with Second Toe Transplant.

OBJECTIVE: To report preoperative planning using 3D printing to plan thumb reconstructions with second toe transplant. METHODS: Between December 2013 and October 2015, the thumbs of five patients with grade 3 thumb defects were reconstructed using a wrap-around flap and second toe transplant aided by 3D printing technology. CT scans of hands and feet were analyzed using Boholo surgical simulator software (www.boholo.com). This allowed for the creation of a mirror image of the healthy thumb using the uninjured thumb. Using 3D images of the reconstructed thumb, a model of the big toe and the second toe was created to understand the dimensions of the donor site. This model was also used to repair the donor site defect by designing appropriate iliac bone and superficial circumflex iliac artery flaps. The polylactic acid model of the donor toes and reconstructed thumb was produced using 3D printing. Surgically, the wrap-around flap of the first dorsal metatarsal artery and vein combined with the joint and bone of the second toe was based upon the model donor site. Sensation was reconstructed by anastomosing the dorsal nerve of the foot and the plantar digital nerve of the great toe. Patients commenced exercises 2 weeks after surgery. RESULTS: All reconstructed thumbs survived, although partial flap necrosis occurred in one case. This was managed with regular dressing changes. Patients were followed up for 3-15 months. The lengths of the reconstructed thumbs are 34-49 mm. The widths of the thumb nail beds are 16-19 mm, and the thickness of the digital pulp is 16-20 mm. The thumb opposition function was 0-1.5 cm; the extension angle was 5°-20° (mean, 16°), and the angle of flexion was 38°-55° (mean, 47°). Two-point discrimination was 9-11 mm (mean, 9.6 mm). The reconstructed thumbs had good appearance, function and sensation. Based on the criteria set forth by the Standard on Approval of Reconstructed Thumb and Finger Functional Assessment of the Chinese Medical Association, the results were considered excellent for four cases and good for one case. The success rate was 100%. CONCLUSIONS: When planning a wrap-around flap and second toe transplant to reconstruct a thumb, both the donor and recipient sites can be modeled using 3D printing. This can shorten the operative time by supplying digital and accurate schematics for the operation. It can also optimize the function and appearance of the reconstructed thumb while minimizing damage to the donor site.

PKC-NF-κB are involved in CCL2-induced Nav1.8 expression and channel function in dorsal root ganglion neurons.

CCL2 [chemokine (C-C motif) ligand 2] contributes to the inflammation-induced neuropathic pain through activating VGSC (voltage-gated sodium channel)-mediated nerve impulse conduction, but the underlying mechanism is currently unknown. Our study aimed to investigate whether PKC (protein kinase C)-NF-κB (nuclear factor κB) is involved in CCL2-induced regulation of voltage-gated sodium Nav1.8 currents and expression. DRG (dorsal root ganglion) neurons were prepared from adult male Sprague-Dawley rats and incubated with various concentration of CCL2 for 24 h. Whole-cell patch-clamps were performed to record the Nav1.8 currents in response to the induction by CCL2. After being pretreated with 5 and10 nM CCL2 for 16 h, CCR2 [chemokine (C-C motif) receptor 2] and Nav1.8 expression significantly increased and the peak currents of Nav1.8 elevated from the baseline 46.53±4.53 pA/pF to 64.28±3.12 pA/pF following 10 nM CCL2 (P<0.05). Compared with the control, significant change in Nav1.8 current density was observed when the CCR2 inhibitor INCB3344 (10 nM) was applied. Furthermore, inhibition of PKC by AEB071 significantly eliminated CCL2-induced elevated Nav1.8 currents. In vitro PKC kinase assays and autoradiograms suggested that Nav1.8 within DRG neurons was a substrate of PKC and direct phosphorylation of the Nav1.8 channel by PKC regulates its function in these neurons. Moreover, p65 expression was significantly higher in CCL2-induced neurons (P<0.05), and was reversed by treatment with INCB3344 and AEB071. PKC-NF-κB are involved in CCL2-induced elevation of Nav1.8 current density by promoting the phosphorylation of Nav1.8 and its expression.

Berberine inhibits fluphenazine-induced up-regulation of CDR1 in Candida albicans.

Over-expression of the Candida drug resistance gene CDR1 is a common mechanism generating azole-resistant Candida albicans in clinical isolates. CDR1 is transcriptionally activated through the binding of the transcription factor Tac1p to the cis-acting drug-responsive element (DRE) in its promoter. We previously demonstrated that the combination of fluconazole (FLC) and berberine (BBR) produced significant synergy when used against FLC-resistant C. albicans in vitro. In this study, we found that BBR inhibited both the up-regulation of CDR1 mRNA and the transport function of Cdr1p induced by fluphenazine (FNZ). Further, electrophoretic mobility shift assays suggested that the transcription activation complex of protein-DRE was disrupted by BBR, and electrospray ionization mass spectrometry analysis showed that BBR bound to the DRE of CDR1. Thus we propose that BBR inhibits the FNZ-induced transcriptional activation of CDR1 in C. albicans by blocking transcription factor binding to the DRE of CDR1. These results contribute to our understanding of the mechanism of synergistic effect of BBR and FLC.

[Derivatization of berberine based on its synergistic antifungal activity with fluconazole against fluconazole-resistant Candida albicans].

Abstract: Our previous work revealed berberine can significantly enhance the susceptibility of fluconazole against fluconazole-resistant Candida albicans, which suggested that berberine has synergistic antifungal activity with fluconazole. Preliminary SAR of berberine needs to be studied for the possibility of investigating its target and SAR, improving its drug-likeness, and exploring new scaffold. In this work, 13-substitutited benzyl berberine derivatives and N-benzyl isoquinoline analogues were synthesized and characterized by 1H NMR and MS. Their synergetic activity with fluconazole against fluconazole-resistant Candida albicans was evaluated in vitro. The 13-substitutited benzyl berberine derivatives 1a-1e exhibited comparable activity to berberine, which suggested that the introduction of functional groups to C-13 can maintain its activity. The N-benzyl isoquinolines, which were designed as analogues of berberine with its D ring opened, exhibited lower activity than berberine. However, compound 2b, 2c, and 4b showed moderate activity, which indicated that berberine may be deconstructed to new scaffold with synergistic antifungal activity with fluconazole. The results of our research may be helpful to the SAR studies on its other biological activities.

[Zoledronic acid incorporated in chitosan/calcium phosphate ceramic: characterization and in vitro response of osteoblast cells].

OBJECTIVE: To develop a new local delivery system, zoledronic-acid-loaded chitosan/calcium phosphate ceramic, and to determine its characterization and in vitro response of osteoblast cells. METHODS: Zoledronic-acid-loaded chitosan/calcium phosphate ceramic were prepared by solution casting method at a concentration of 10(-5), 10(-4), and 10(-3) mol/L, respectively. The physicochemical properties of the resulting materials were determined using SEM and FTIR. Drug absorbance was measured using CCK-8 colorimetric assay and alkaline phosphatase assay to detect the effect of drug-loaded materials on the proliferation and differentiation of osteoblasts. RESULTS: After ZOL loading, SEM showed that porous calcium phosphate ceramic was coated with chitosan evenly. The IR spectra indicated that drug absorption peaks were shifted and a new one was formed for the drug-loaded biomaterials. The material at the highest concentration could inhibit the proliferation and alkaline phosphatase activities of osteoblast cells, but no such effect was found at a drug-loading concentration of 10(-4)-10(-5) mol/L. CONCLUSION: We confirmed that the local delivery system in this study has ability of loading ZOL. The biomaterial with high drug concentrations inhibits the proliferation and differentiation of osteoblasts, but not when the drug concentrations are low.

[Preparation of zoledronic-acid-loaded collagen membrane].

OBJECTIVE: To develop a new local delivery system, osteoclastic-inhibitor-loaded collagen membrane, and to evaluate its drug loading and drug release properties. METHODS: Efforts were made to develop the drug-loaded membranes by combining two commercially available collagen barrier membranes (Bio-Gide and BME-10X) with zoledronic acid (ZA). The physicochemical and pharmacological properties of resulting materials were determined using SEM, EDS, FTIR, and HPLC. RESULTS: After ZA loading, the micropores between the thin collagen fibers in the Bio-Gide disappeared, whereas crystalloid powders appeared on the surface of pore walls in BME-10X. Phosphorus was detected on both drug-loaded membranes. The Amides shifted. With the same drug solution, Bio-Gide presented larger amount of ZA loading and slower ZA release than BME-10X. ZA loading did not affect the 3D fiber network and the degradation of membranes. CONCLUSION: Both collagen membranes load ZA successfully and delay drug release. But Bio-Gide shows higher loading values and slower release than BME-10X.